The calcium-binding EF-hand proteins S100A8 and S100A9 are released from damaged keratinocytes as the heterodimeric protein calprotectin. Calprotectin stimulates keratinocyte activation, characterized by proliferation, migration, and secretion of pro-inflammatory cytokines, as well as neutrophil chemotaxis. Like calprotectin, the Slug zinc finger transcription factor also is damage-inducible and promotes keratinocyte activation. In the skin, both calprotectin and Slug play important roles in wound healing, carcinogenesis, and inflammation. Preliminary findings indicate that calprotectin induction in keratinocytes is Slug-dependent and suggest that calprotectin may be responsible, at least in part, for mediating Slug-stimulated keratinocyte activation. Moreover, calprotectin is produced by both keratinocytes and leukocytes and can act via autocrine and paracrine routes to stimulate keratinocyte and leukocyte activation and the secretion of additional calprotectin. The novel hypothesis to be tested in this application is that Slug and calprotectin participate in a potent feed-forward mechanism of keratinocyte activation. This hypothesis will be tested using in vivo and in vitro models to 1) determine the extent to which calprotectin induction is dependent upon Slug and identify the pathway(s) by which Slug modulates expression of calprotectin; 2) determine the extent to which calprotectin mediates Slug-dependent keratinocyte activation in the skin and identify the cell surface receptor(s) that stimulates keratinocyte activation in response to calprotectin; and 3) determine the contribution of leukocyte- derived calprotectin to keratinocyte activation. Based on previous studies, the NFB pathway is a likely candidate for Slug modulation of calprotectin production, and RAGE is the probable keratinocyte receptor for calprotectin. The contribution of these putative keratinocyte activation pathway components will be examined using specific inhibitors, but the participation of other signaling molecules will also be considered and tested as appropriate. Involvement of leukocyte-derived versus keratinocyte-derived calprotectin in keratinocyte activation will be tested in bone marrow chimeric mice that express calprotectin only in keratinocytes or leukocytes. The unexpected association between Slug induction and calprotectin secretion has important implications for a wide variety of cutaneous diseases, as it establishes a direct link between cutaneous inflammation mediated by calprotectin and Slug-induced epithelial-mesenchymal transformation important in wound healing and tumor progression. It may be possible to take advantage of the interaction between Slug and calprotectin to design new strategies to prevent and treat a wide spectrum of skin diseases. PUBLIC HEALTH RELEVANCE: The skin is subject to many environmental insults, including wounds, sunlight, and toxic substances. We are studying the molecular pathways by which the skin responds to these insults with the aim of being able to enhance desirable responses like wound healing and to mitigate undesirable responses like sunburn. Our studies focus on two components of the pathway and their relationship: Slug and calprotectin, both of which enhance cell migration during wound healing, but also stimulate inflammation during sunburn.